Cardiovascular prostheses

ABSTRACT

Cardiovascular prostheses for treating, reconstructing and replacing damaged or diseased cardiovascular tissue that are formed from acellular extracellular matrix (ECM). The cardiovascular prostheses comprise various compositions, such as ECM based compositions, and structures, such as particulate structures, mesh constructs, encasement structures, coated structures and multi-sheet laminate structures.

CROSS-REFERENCES TO RELATED APPLICATIONS

This application is a continuation-in-part of U.S. application Ser. No.13/328,287, filed on Dec. 16, 2011, which claims the benefit of U.S.Provisional Application No. 61/425,287, filed on Dec. 20, 2010.

FIELD OF THE INVENTION

The present invention relates to methods and apparatus for treatingdamaged or diseased cardiovascular structures. More particularly, thepresent invention relates to cardiovascular prostheses for treatingand/or reconstructing damaged or diseased cardiovascular structures.

BACKGROUND OF THE INVENTION

As is well known in the art, various cardiovascular prostheses are oftenemployed to treat and reconstruct damaged or diseased cardiovascularstructures and associated tissue, such as cardiovascular vessels andheart tissue. However, despite the growing sophistication of medicaltechnology, the use of prostheses to treat or replace damaged biologicaltissue remains a frequent and serious problem in health care. Theproblem is often associated with the materials employed to construct theprostheses.

As is also well known in the art, the optimal prostheses material shouldbe chemically inert, non-carcinogenic, capable of resisting mechanicalstress, capable of being fabricated in the form required andsterilizable. Further, the material should be resistant to physicalmodification by tissue fluids, and not excite an inflammatory reaction,induce a state of allergy or hypersensitivity, or, in some cases,promote visceral adhesions.

Various materials and/or structures have thus been employed to constructprostheses that satisfy the aforementioned optimal characteristics. Suchmaterials and structures include tantalum gauze, stainless mesh,Dacron®, Orlon®, Fortisan®, nylon, knitted polypropylene (e.g.,Marlex®), microporous expanded-polytetrafluoroethylene (e.g.,Gore-Tex®), Dacron reinforced silicone rubber (e.g., Silastic®),polyglactin 910 (e.g., Vicryl®), polyester (e.g., Mersilene®),polyglycolic acid (e.g., Dexon®), processed sheep dermal collagen,crosslinked bovine pericardium (e.g., Peri-Guard®), and preserved humandura (e.g., Lyodura®).

As discussed in detail below, although some of the noted prosthesismaterials satisfy some of the aforementioned optimal characteristics,few, if any, satisfy all of the optimal characteristics.

Metallic mesh structures, e.g., stainless steel meshes, are generallyinert and resistant to infection. Metallic mesh structures are, however,prone to fragmentation, which can, and in many instances will, occurafter the first year of administration.

Synthetic mesh structures are easily molded and, except for nylon,retain their tensile strength in or on the body. Synthetic meshstructures are, however, typically non-resorbable and susceptibility toinfection.

A major problem associated with Marlex®, i.e. polypropylene, meshstructures is that with scar contracture, polypropylene mesh structuresbecome distorted and separate from surrounding normal tissue.

A major problem associated with Gore-Tex®, i.e. polytetrafluoroethylene,mesh structures is that in a contaminated wound it does not allow forany macromolecular drainage, which limits treatment of infections.

Mammalian tissue, such as extracellular matrix (ECM), is also oftenemployed to construct cardiovascular prostheses. Illustrative are theprostheses disclosed in U.S. Pat. Nos. 3,562,820 and 4,902,508. FurtherECM prostheses (i.e. multi-sheet laminate structures) are disclosed inU.S. Pat. No. 8,808,363 and Applicant's Co-Pending application Ser. Nos.14/031,423, 14/337,915, 14/566,155 and 14/566,306, which areincorporated by reference herein in their entirety.

Although many of the ECM based cardiovascular prostheses satisfy many ofthe aforementioned optimal characteristics, when the ECM graft comprisestwo or more sheets, i.e. a multi-sheet laminate, such as disclosed inCo-pending application Ser. No. 14/031,423, the laminate structures can,and in some instances will, delaminate.

Thus, readily available, versatile cardiovascular prostheses that arenot prone to calcification, thrombosis, intimal hyperplasia anddelamination would fill a substantial and growing clinical need.

It is therefore an object of the present invention to providecardiovascular prostheses that substantially reduce or eliminate (i) therisk of thrombosis, (ii) intimal hyperplasia after intervention in avessel, (iii) the harsh biological responses associated withconventional polymeric and metal prostheses, and (iv) the formation ofbiofilm, inflammation and infection.

It is another object of the present invention to provide cardiovascularprostheses that modulate inflammation and induce host tissueproliferation, remodeling and regeneration of new tissue and tissuestructures with site-specific structural and functional properties whendelivered to damaged cardiovascular tissue.

It is another object of the present invention to provide cardiovascularprostheses that are capable of administering a pharmacological agent tohost tissue and, thereby produce a desired biological and/or therapeuticeffect.

SUMMARY OF THE INVENTION

The present invention is directed to biodegradable and remodelablecardiovascular prostheses for treating, reconstructing or replacingdamaged or diseased cardiovascular structures and associated tissue.

According to the invention, the cardiovascular prostheses can comprisevarious structures and compositions, including, but not limited to,particulate structures, mesh constructs, encasement structures, coatedstructures and multi-sheet laminate structures.

In a preferred embodiment of the invention, when the cardiovascularprostheses are disposed proximate (i.e. delivered or administered to)damaged tissue, the cardiovascular prostheses induce neovascularizationand/or remodeling of the damaged tissue, without inducing an adverseinflammatory response.

In some embodiments of the invention, when the cardiovascular prosthesesare disposed proximate damaged tissue, the cardiovascular prosthesesmodulate inflammation of the damaged tissue and, induceneovascularization, host cell and tissue proliferation, and regenerationof new tissue and tissue structures.

In some embodiments, the cardiovascular prostheses comprise an ECMcomposition comprising acellular ECM derived from a mammalian tissuesource.

According to the invention, the mammalian tissue sources can comprise,without limitation, small intestine submucosa (SIS), urinary bladdersubmucosa (UBS), urinary basement membrane (UBM), liver basementmembrane (LBM), stomach submucosa (SS), mesothelial tissue, placentaltissue and cardiac tissue, pericardial tissue.

In some embodiments of the invention, the ECM composition comprises atleast one additional biologically active agent or composition, i.e. anagent that induces or modulates a physiological or biological process,or cellular activity, e.g., induces proliferation, and/or growth and/orregeneration of tissue.

In some embodiments, the biologically active agent comprises a growthfactor, such as, without limitation, a transforming growth factor-alpha(TGF-α), transforming growth factor-beta (TGF-β), basic fibroblastgrowth factor (bFGF) and vascular epithelial growth factor (VEGF).

In some embodiments, the ECM composition comprises at least onepharmacological agent or composition (or drug), i.e. an agent orcomposition that is capable of producing a desired biological effect invivo, e.g., stimulation or suppression of apoptosis, stimulation orsuppression of an immune response, etc.

Suitable pharmacological agents and compositions include, withoutlimitation, antibiotics, anti-fibrotics, anti-viral agents, analgesics,anti-inflammatories, anti-neoplastics, anti-spasmodics, andanti-coagulants and/or anti-thrombotic agents.

In some embodiments of the invention, the pharmacological agentcomprises a statin, i.e. a HMG-CoA reductase inhibitor, such ascerivastatin.

In some embodiments of the invention, the pharmacological agentcomprises an antibiotic, such as vancomycin and gentamicin.

In some embodiments of the invention, the pharmacological agentcomprises an antimicrobial, such as silver particles and copperparticles.

In some embodiments, the cardiovascular prostheses comprise anECM-mimicking composition comprising poly(glycerol sebacate) (PGS).

In some embodiments, the ECM-mimicking composition further comprises atleast one of the aforementioned biologically active agents and/orpharmacological agents.

In some embodiments, the cardiovascular prostheses comprise anECM/ECM-mimicking composition comprising acellular ECM and PGS.

In some embodiments, the ECM/ECM-mimicking composition further comprisesat least one of the aforementioned biologically active agents and/orpharmacological agents.

In some embodiments, the cardiovascular prostheses comprise multi-layerstructures comprising different compositions.

BRIEF DESCRIPTION OF THE DRAWINGS

Further features and advantages will become apparent from the followingand more particular description of the preferred embodiments of theinvention, as illustrated in the accompanying drawings, and in whichlike referenced characters generally refer to the same parts or elementsthroughout the views, and in which:

FIG. 1 is a schematic illustration showing the effects of statins onvascular cell walls, in accordance with the invention;

FIG. 2 is a perspective view of one embodiment of a prosthesis sheetmember, in accordance with the invention;

FIG. 3 is front plan view of the prosthesis sheet member shown in FIG.2, in accordance with the invention;

FIG. 4 is a perspective view of another embodiment of a prosthesis sheetmember, in accordance with the invention;

FIG. 5 is front plan view of the prosthesis sheet member shown in FIG.4, in accordance with the invention;

FIG. 6 is a front plan view of one embodiment of a multi-sheetprosthesis structure in a pre-lamination configuration, in accordancewith the invention;

FIG. 7 is a front plan view of the multi-sheet prosthesis structureshown in FIG. 6 in a laminated configuration, in accordance with theinvention;

FIG. 8 is a perspective view of another embodiment of a multi-sheetprosthesis structure in a pre-laminated configuration, in accordancewith the invention;

FIG. 9 is a front plan view of the multi-sheet prosthesis structureshown in FIG. 8 in a laminated configuration, in accordance with theinvention;

FIG. 10 is a perspective view one embodiment of a prosthesis encasementstructure, in accordance with the invention;

FIG. 11 is a front plan view of the prosthesis encasement structureshown in FIG. 10, in accordance with the invention;

FIG. 12 is a perspective view of another embodiment of a prosthesisencasement structure, in accordance with the invention;

FIG. 13 is a perspective view of one embodiment of a prosthetic meshstructure, in accordance with the invention;

FIG. 14 is a front perspective view of one embodiment of a prostheticparticulate structure, in accordance with the invention;

FIG. 15 is a graphical illustration of in vivo C—C chemokine receptortype 2 (CCR2) expression as a function of time for a cerivastatinaugmented ECM prosthesis, in accordance with the invention;

FIG. 16 is a graphical illustration of in vivo monocyte chemoattractantprotein-1 (MCP-1) expression as a function of time for a cerivastatinaugmented ECM prosthesis, in accordance with the invention;

FIG. 17 is a graphical illustration of lipopolysaccharide (LPS) inducedexpression of MCP-1 in a human monocytic cell line; particularly, THP-1cells, as a function of time for a non-cerivastatin augmented ECMprosthesis, in accordance with the invention;

FIG. 18 is a graphical illustration of LPS induced expression of MCP-1in THP-1 cells as a function of time for a cerivastatin augmented ECMprosthesis, in accordance with the invention; and

FIG. 19 is a graphical illustration of LPS induced expression of MCP-1in THP-1 cells as a function of time for cerivastatin augmented ECMprostheses having various concentrations of cerivastatin, in accordancewith the invention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

Before describing the present invention in detail, it is to beunderstood that this invention is not limited to particularlyexemplified compositions, structures, apparatus, and methods, as suchmay, of course, vary. Thus, although a number of compositions,structures, apparatus, and methods similar or equivalent to thosedescribed herein can be used in the practice of the present invention,the preferred compositions, structures, apparatus, and methods aredescribed herein.

It is also to be understood that the terminology used herein is for thepurpose of describing particular embodiments of the invention only andis not intended to be limiting.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one having ordinaryskill in the art to which the invention pertains.

Further, all publications, patents and patent applications cited herein,whether supra or infra, are hereby incorporated by reference herein intheir entirety.

As used in this specification and the appended claims, the singularforms “a, “an” and “the” include plural referents unless the contentclearly dictates otherwise. Thus, for example, reference to “apharmacological agent” includes two or more such agents and the like.

Further, ranges can be expressed herein as from “about” or“approximately” one particular value, and/or to “about” or“approximately” another particular value. When such a range isexpressed, another embodiment includes from the one particular valueand/or to the other particular value. Similarly, when values areexpressed as approximations, by use of the antecedent “about” or“approximately”, it will be understood that the particular value formsanother embodiment. It will be further understood that the endpoints ofeach of the ranges are significant both in relation to the otherendpoint, and independently of the other endpoint.

Definitions

The terms “extracellular matrix” and “ECM” are used interchangeablyherein, and mean and include a collagen-rich substance that is found inbetween cells in mammalian tissue, and any material processed therefrom,e.g. acellular ECM derived from mammalian tissue sources.

According to the invention, ECM can be derived from a variety ofmammalian tissue sources, including, without limitation, small intestinesubmucosa (SIS), urinary bladder submucosa (UBS), stomach submucosa(SS), central nervous system tissue and epithelium of mesodermal origin,i.e. mesothelial tissue.

The terms “urinary bladder submucosa (UBS)”, “small intestine submucosa(SIS)” and “stomach submucosa (SS)” also mean and include any UBS and/orSIS and/or SS material that includes the tunica mucosa (which includesthe transitional epithelial layer and the tunica propria), submucosallayer, one or more layers of muscularis, and adventitia (a looseconnective tissue layer) associated therewith.

ECM can also be derived from basement membrane of mammaliantissue/organs, including, without limitation, urinary basement membrane(UBM), liver basement membrane (LBM), and amnion, chorion, allograftpericardium, allograft dermis, amniotic membrane, Wharton's jelly,umbilical cord, and combinations thereof.

Additional sources of mammalian basement membrane include, withoutlimitation, spleen tissue, lymph node tissue, salivary gland tissue,prostate tissue, pancreas tissue and tissue from other secreting glands.

The ECM can also be derived from dermal tissue, subcutaneous tissue,placental tissue; cardiac tissue, e.g., pericardial and/or myocardialtissue, kidney tissue, lung tissue, gastrointestinal tissue, i.e. largeand small intestinal, appendix, omentum and pancreas tissue, andcombinations thereof.

ECM can also be derived from other sources, including, withoutlimitation, collagen from plant sources and synthesized extracellularmatrices, i.e. cell cultures. ECM can also comprise ECM synthesized invitro, e.g., collagen producing cell lines, and collagen and ECM fromnon-mammalian tissue sources, such as, without limitation, avian,reptilian, fish, and other marine sources.

The terms “decellularized” and “acellular” are used interchangeablyherein in connection with ECM, and mean and include ECM derived frommammalian tissue subjected to a decellularized process and, hence,exhibits a reduced glycosaminoglycan (GAG) content and markedly alteredcollagen and fibronectin structures compared to naturally occurringmammalian tissue.

The term “medical device”, as used herein, means and includes atherapeutic, surgical or prosthetic device configured to modulate abiological function of a warm blooded mammal, including humans andprimates; avians; domestic household or farm animals, such as cats,dogs, sheep, goats, cattle, horses and pigs; laboratory animals, such asmice, rats and guinea pigs; fish; reptiles; zoo and wild animals; andthe like. The term “medical device” thus includes, without limitation,an implantable medical device, such as a pacemaker, defibrillator,synthetic heart valve, ventricular assist device, artificial heart,physiological sensor, catheter and associated components thereof,including electrical leads and lines associated therewith.

The term “angiogenesis”, as used herein, means a physiologic processinvolving the growth of new blood vessels from pre-existing bloodvessels.

The term “neovascularization”, as used herein, means and includes theformation of functional vascular networks that can be perfused by bloodor blood components. Neovascularization includes angiogenesis, buddingangiogenesis, intussuceptive angiogenesis, sprouting angiogenesis,therapeutic angiogenesis and vasculogenesis.

The term “adverse inflammatory response”, as used herein, means andincludes a physiological response that is sufficient to induceconstitutive clinically relevant expression of pro-inflammatorycytokines, such as interleukin-1 beta (IL-1β) and monocytechemoattractant protein-1 (MCP-1) in vivo.

The term “adverse biological response”, as used herein, means andincludes a physiological response that is sufficient to induce abiological process and/or restrict a phase associated with biologicaltissue healing in vivo, including without limitation, neovascularizationand remodeling of the damaged biological tissue. The term “adversebiological response” thus includes an “adverse inflammatory response”,e.g. development of fibrotic tissue.

The terms “biologically active agent” and “biologically activecomposition” are used interchangeably herein, and mean and include agentor composition that induces or modulates a physiological or biologicalprocess, or cellular activity, e.g., induces proliferation, and/orgrowth and/or regeneration of tissue.

The terms “biologically active agent” and “biologically activecomposition” thus mean and include, without limitation, the followinggrowth factors and compositions comprising same: platelet derived growthfactor (PDGF), epidermal growth factor (EGF), transforming growth factoralpha (TGF-α), transforming growth factor beta (TGF-β), basic fibroblastgrowth factor (bFGF) (also referred to as fibroblast growth factor-2(FGF-2)), vascular epithelial growth factor (VEGF), hepatocyte growthfactor (HGF), insulin-like growth factor (IGF), nerve growth factor(NGF), platelet derived growth factor (PDGF), tumor necrosis factoralpha (TNF-α), and placental growth factor (PLGF).

The terms “biologically active agent” and “biologically activecomposition” also mean and include, without limitation, the followingcells and compositions comprising same: human embryonic stem cells,fetal cardiomyocytes, myofibroblasts, mesenchymal stem cells,autotransplated expanded cardiomyocytes, adipocytes, totipotent cells,pluripotent cells, blood stem cells, myoblasts, adult stem cells, bonemarrow cells, mesenchymal cells, embryonic stem cells, parenchymalcells, epithelial cells, endothelial cells, mesothelial cells,fibroblasts, osteoblasts, chondrocytes, exogenous cells, endogenouscells, stem cells, hematopoietic stem cells, bone-marrow derivedprogenitor cells, myocardial cells, skeletal cells, fetal cells,undifferentiated cells, multi-potent progenitor cells, unipotentprogenitor cells, monocytes, cardiac myoblasts, skeletal myoblasts,macrophages, capillary endothelial cells, xenogeneic cells, allogeneiccells, and post-natal stem cells.

The terms “biologically active agent” and “biologically activecomposition” also mean and include, without limitation, the followingbiologically active agents (referred to interchangeably herein as a“protein”, “peptide” and “polypeptide”) and compositions comprisingsame: collagen (types I-V), proteoglycans, glycosaminoglycans (GAGs),glycoproteins, growth factors, cytokines, cell-surface associatedproteins, cell adhesion molecules (CAM), angiogenic growth factors,endothelial ligands, matrikines, cadherins, immunoglobins, fibrilcollagens, non-fibrillar collagens, basement membrane collagens,multiplexins, small-leucine rich proteoglycans, decorins, biglycans,fibromodulins, keratocans, lumicans, epiphycans, heparin sulfateproteoglycans, perlecans, agrins, testicans, syndecans, glypicans,serglycins, selectins, lecticans, aggrecans, versicans, neurocans,brevicans, cytoplasmic domain-44 (CD-44), macrophage stimulatingfactors, amyloid precursor proteins, heparins, chondroitin sulfate B(dermatan sulfate), chondroitin sulfate A, heparin sulfates, hyaluronicacids, fibronectins, tenascins, elastins, fibrillins, laminins,nidogen/enactins, fibulin I, fibulin II, integrins, transmembranemolecules, thrombospondins, osteopontins, and angiotensin convertingenzymes (ACE).

The terms “biologically active agent” and “biologically activecomposition” also mean and include an “exosome”, “microsome” or“micro-vesicle,” which are used interchangeably herein, and mean andinclude a micellar body formed from a hydrocarbon monolayer or bilayerconfigured to contain or encase a composition of matter, such as abiologically active agent. The terms “exosome”, “microsome” and“micro-vesicle” thus include, without limitation, a micellar body formedfrom a lipid layer configured to contain or encase biologically activeagents and/or combinations thereof.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” are used interchangeably herein, and mean and includean agent, drug, compound, composition of matter or mixture thereof,including its formulation, which provides some therapeutic, oftenbeneficial, effect. This includes any physiologically orpharmacologically active substance that produces a localized or systemiceffect or effects in animals, including warm blooded mammals, humans andprimates; avians; domestic household or farm animals, such as cats,dogs, sheep, goats, cattle, horses and pigs; laboratory animals, such asmice, rats and guinea pigs; fish; reptiles; zoo and wild animals; andthe like.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” thus mean and include, without limitation,antibiotics, anti-fibrotics, antiarrhythmic agents, anti-viral agents,analgesics, steroidal anti-inflammatories, non-steroidalanti-inflammatories, anti-neoplastics, anti-spasmodics, modulators ofcell-extracellular matrix interactions, proteins, hormones, growthfactors, matrix metalloproteinases (MMPs), enzymes and enzymeinhibitors, anticoagulants and/or anti-thrombotic agents, DNA, RNA,modified DNA and RNA, NSAIDs, inhibitors of DNA, RNA or proteinsynthesis, polypeptides, oligonucleotides, polynucleotides,nucleoproteins, compounds modulating cell migration, compoundsmodulating proliferation and growth of tissue, and vasodilating agents.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” thus include, without limitation, atropine,tropicamide, dexamethasone, dexamethasone phosphate, betamethasone,betamethasone phosphate, prednisolone, triamcinolone, triamcinoloneacetonide, fluocinolone acetonide, anecortave acetate, budesonide,cyclosporine, FK-506, rapamycin, ruboxistaurin, midostaurin,flurbiprofen, suprofen, ketoprofen, diclofenac, ketorolac, nepafenac,lidocaine, neomycin, polymyxin b, bacitracin, gramicidin, gentamicin,oyxtetracycline, ciprofloxacin, ofloxacin, tobramycin, amikacin,vancomycin, cefazolin, ticarcillin, chloramphenicol, miconazole,itraconazole, trifluridine, vidarabine, ganciclovir, acyclovir,cidofovir, ara-amp, foscarnet, idoxuridine, adefovir dipivoxil,methotrexate, carboplatin, phenylephrine, epinephrine, dipivefrin,timolol, 6-hydroxydopamine, betaxolol, pilocarpine, carbachol,physostigmine, demecarium, dorzolamide, brinzolamide, latanoprost,sodium hyaluronate, insulin, verteporfin, pegaptanib, ranibizumab, andother antibodies, antineoplastics, anti-VEGFs, ciliary neurotrophicfactor, brain-derived neurotrophic factor, bFGF, Caspase-1 inhibitors,Caspase-3 inhibitors, α-Adrenoceptors agonists, NMDA antagonists, Glialcell line-derived neurotrophic factors (GDNF), pigmentepithelium-derived factor (PEDF), and NT-3, NT-4, NGF, IGF-2.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” further mean and include the following Class I-ClassV antiarrhythmic agents: (Class Ia) quinidine, procainamide anddisopyramide; (Class Ib) lidocaine, phenytoin and mexiletine; (Class Ic)flecainide, propafenone and moricizine; (Class II) propranolol, esmolol,timolol, metoprolol and atenolol; (Class III) amiodarone, sotalol,ibutilide and dofetilide; (Class IV) verapamil and diltiazem) and (ClassV) adenosine and digoxin.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” further mean and include, without limitation, thefollowing antimicrobials: silver particles, copper particles, cobaltparticles, nickel particles, zinc particles, zirconium particles,molybdenum particles, lead particles and mixtures thereof.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” further mean and include, without limitation, thefollowing antibiotics: aminoglycosides, cephalosporins, chloramphenicol,clindamycin, erythromycins, fluoroquinolones, macrolides, azolides,metronidazole, penicillins, tetracyclines, trimethoprim-sulfamethoxazoleand vancomycin.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” further mean and include, without limitation, thefollowing anti-fibrotics: paclitaxel, sirolimus and derivatives thereof,including everolimus.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” further include, without limitation, the followingsteroids: andranes (e.g., testosterone), cholestanes, cholic acids,corticosteroids (e.g., dexamethasone), estraenes (e.g., estradiol) andpregnanes (e.g., progesterone).

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” can further include one or more classes of narcoticanalgesics, including, without limitation, morphine, codeine, heroin,hydromorphone, levorphanol, meperidine, methadone, oxycodone,propoxyphene, fentanyl, methadone, naloxone, buprenorphine, butorphanol,nalbuphine and pentazocine.

The terms “pharmacological agent”, “active agent”, “drug” and “activeagent formulation” can further include one or more classes of topical orlocal anesthetics, including, without limitation, esters, such asbenzocaine, chloroprocaine, cocaine, cyclomethycaine,dimethocaine/larocaine, piperocaine, propoxycaine, procaine/novacaine,proparacaine, and tetracaine/amethocaine. Local anesthetics can alsoinclude, without limitation, amides, such as articaine, bupivacaine,cinchocaine/dibucaine, etidocaine, levobupivacaine,lidocaine/lignocaine, mepivacaine, prilocaine, ropivacaine, andtrimecaine. Local anesthetics can further include combinations of theabove from either amides or esters.

The terms “anti-inflammatory” and “anti-inflammatory agent” are alsoused interchangeably herein, and mean and include a “pharmacologicalagent” and/or “active agent formulation”, which, when a therapeuticallyeffective amount is administered to a subject, prevents or treats bodilytissue inflammation i.e. the protective tissue response to injury ordestruction of tissues, which serves to destroy, dilute, or wall offboth the injurious agent and the injured tissues.

Anti-inflammatory agents thus include, without limitation, alclofenac,alclometasone dipropionate, algestone acetonide, alpha amylase,amcinafal, amcinafide, amfenac sodium, amiprilose hydrochloride,anakinra, anirolac, anitrazafen, apazone, balsalazide disodium,bendazac, benoxaprofen, benzydamine hydrochloride, bromelains,broperamole, budesonide, carprofen, cicloprofen, cintazone, cliprofen,clobetasol propionate, clobetasone butyrate, clopirac, cloticasonepropionate, cormethasone acetate, cortodoxone, decanoate, deflazacort,delatestryl, depo-testosterone, desonide, desoximetasone, dexamethasonedipropionate, diclofenac potassium, diclofenac sodium, diflorasonediacetate, diflumidone sodium, diflunisal, difluprednate, diftalone,dimethyl sulfoxide, drocinonide, endrysone, enlimomab, enolicam sodium,epirizole, etodolac, etofenamate, felbinac, fenamole, fenbufen,fenclofenac, fenclorac, fendosal, fenpipalone, fentiazac, flazalone,fluazacort, flufenamic acid, flumizole, flunisolide acetate, flunixin,flunixin meglumine, fluocortin butyl, fluorometholone acetate,fluquazone, flurbiprofen, fluretofen, fluticasone propionate,furaprofen, furobufen, halcinonide, halobetasol propionate, halopredoneacetate, ibufenac, ibuprofen, ibuprofen aluminum, ibuprofen piconol,ilonidap, indomethacin, indomethacin sodium, indoprofen, indoxole,intrazole, isoflupredone acetate, isoxepac, isoxicam, ketoprofen,lofemizole hydrochloride, lomoxicam, loteprednol etabonate,meclofenamate sodium, meclofenamic acid, meclorisone dibutyrate,mefenamic acid, mesalamine, meseclazone, mesterolone,methandrostenolone, methenolone, methenolone acetate, methylprednisolonesuleptanate, momiflumate, nabumetone, nandrolone, naproxen, naproxensodium, naproxol, nimazone, olsalazine sodium, orgotein, orpanoxin,oxandrolane, oxaprozin, oxyphenbutazone, oxymetholone, paranylinehydrochloride, pentosan polysulfate sodium, phenbutazone sodiumglycerate, pirfenidone, piroxicam, piroxicam cinnamate, piroxicamolamine, pirprofen, prednazate, prifelone, prodolic acid, proquazone,proxazole, proxazole citrate, rimexolone, romazarit, salcolex,salnacedin, salsalate, sanguinarium chloride, seclazone, sennetacin,stanozolol, sudoxicam, sulindac, suprofen, talmetacin, talniflumate,talosalate, tebufelone, tenidap, tenidap sodium, tenoxicam, tesicam,tesimide, testosterone, testosterone blends, tetrydamine, tiopinac,tixocortol pivalate, tolmetin, tolmetin sodium, triclonide,triflumidate, zidometacin, and zomepirac sodium.

The term “pharmacological composition”, as used herein, means andincludes a composition comprising a “pharmacological agent” and/or a“biologically active agent” and/or any additional agent or componentidentified herein.

The term “therapeutically effective”, as used herein, means that theamount of the “pharmacological agent” and/or “biologically active agent”and/or “pharmacological composition” administered is of sufficientquantity to ameliorate one or more causes, symptoms, or sequelae of adisease or disorder. Such amelioration only requires a reduction oralteration, not necessarily elimination, of the cause, symptom, orsequelae of a disease or disorder.

The term “adolescent”, as used herein, means and includes a mammal thatis preferably less than three (3) years of age.

The terms “patient” and “subject” are used interchangeably herein, andmean and include warm blooded mammals, humans and primates; avians;domestic household or farm animals, such as cats, dogs, sheep, goats,cattle, horses and pigs; laboratory animals, such as mice, rats andguinea pigs; fish; reptiles; zoo and wild animals; and the like.

The term “comprise” and variations of the term, such as “comprising” and“comprises,” means “including, but not limited to” and is not intendedto exclude, for example, other additives, components, integers or steps.

The following disclosure is provided to further explain in an enablingfashion the best modes of performing one or more embodiments of thepresent invention. The disclosure is further offered to enhance anunderstanding and appreciation for the inventive principles andadvantages thereof, rather than to limit in any manner the invention.The invention is defined solely by the appended claims including anyamendments made during the pendency of this application and allequivalents of those claims as issued.

The present invention is directed to resilient, non-antigenic,biodegradable, remodelable (or bioremodelable) and, hence,biocompatible, cardiovascular prostheses that can be used to repair,augment, or replace mammalian tissues and organs.

As indicated above and discussed in detail below, in a preferredembodiment of the invention, when the cardiovascular prostheses aredisposed proximate (i.e. delivered or administered to) damaged tissue,the cardiovascular prostheses induce neovascularization and/orremodeling of the damaged tissue, without inducing an adverseinflammatory response.

More preferably, when the cardiovascular prostheses are disposedproximate damaged tissue, the cardiovascular prostheses modulateinflammation of the damaged tissue and, induce neovascularization, hostcell and tissue proliferation, and regeneration of new tissue and tissuestructures.

The cardiovascular prostheses can thus be employed to treat variousdisorders, including, without limitation, atrial fibrillation (pre- andpost-operative) and other causes of ventricular arrhythmias and the rootcauses thereof, damaged or diseased biological tissue, including,without limitation, cardiovascular tissue, e.g., infarct tissue, anddamaged and diseased mammalian organs and structures, including, withoutlimitation, cardiac vessels and valves, such as bicuspid, tricuspid andpulmonary valves, myocardium, pericardium, arteries, veins, trachea,esophagus, etc.

As indicated above, the cardiovascular prostheses can comprise variouscompositions and structures, including, but not limited to, particulatestructures, mesh constructs, encasement structures, coated structuresand multi-sheet laminate structures.

In some embodiments, the cardiovascular prostheses comprise an ECMcomposition comprising acellular ECM derived from a mammalian tissuesource.

According to the invention, the mammalian tissue sources can comprise,without limitation, small intestine tissue, large intestine tissue,stomach tissue, lung tissue, liver tissue, kidney tissue, pancreastissue, placental tissue, cardiac tissue, bladder tissue, prostatetissue, tissue surrounding growing enamel, tissue surrounding growingbone, and any fetal tissue from any mammalian organ.

In some embodiments of the invention, the mammalian tissue sourcescomprise, small intestine submucosa (SIS), urinary bladder submucosa(UBS), urinary basement membrane (UBM), liver basement membrane (LBM),stomach submucosa (SS), mesothelial tissue, placental tissue and cardiactissue.

According to the invention, the ECM composition can comprise acellularECM derived from one (1) mammalian tissue source or acellular ECMderived from different mammalian tissue sources.

In a preferred embodiment, the mammalian tissue source comprises anadolescent mammalian tissue source, i.e. an adolescent mammal, such as apiglet, which is preferably less than three (3) years of age.

According to the invention, an ECM material can be decellularized toprovide acellular ECM by various conventional means.

According to the invention, the ECM material can be decellularized viaone of the conventional decellularization methods disclosed in U.S. Pat.Nos. 7,550,004, 7,244,444, 6,379,710, 6,358,284, 6,206,931, 5,733,337and 4,902,508 and U.S. application Ser. No. 12/707,427; which areincorporated by reference herein in their entirety.

In some embodiments of the invention, the ECM material is decellularizedvia one of the unique Novasterilis™ processes disclosed in U.S. Pat. No.7,108,832 and U.S. patent application Ser. No. 13/480,204; which areincorporated by reference herein in their entirety.

As stated above, in some embodiments of the invention, the ECMcomposition comprises at least one additional or supplementalbiologically active agent or composition, i.e. an agent that induces ormodulates a physiological or biological process, or cellular activity,e.g., induces proliferation, and/or growth and/or regeneration oftissue.

In a preferred embodiment of the invention, the supplementalbiologically active agent is similarly derived from an adolescentmammal, i.e. a mammal less than three (3) years of age.

Suitable supplemental biologically active agents include any of theaforementioned biologically active agents, including, withoutlimitation, the aforementioned cells and proteins.

In some embodiments, the supplemental biologically active agentcomprises a growth factor, such as, without limitation, a transforminggrowth factor-alpha (TGF-α), transforming growth factor-beta (TGF-β),basic fibroblast growth factor (bFGF) and vascular epithelial growthfactor (VEGF).

In some embodiments, the wt. % of the supplemental biologically activeagent in the ECM composition (and ECM-mimicking, ECM/ECM-mimicking, andstatin augmented compositions of the invention, discussed in detailbelow) is in the range of approximately 0.0001-20 wt. %, morepreferably, in the range of approximately 0.001-1 wt.

In a preferred embodiment, the wt. % of the biologically active agent inthe ECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is sufficient to induce ormodulate a physiological or biological process, without inducing anadverse biological response, e.g., a physiological response that issufficient to induce constitutive clinically relevant expression ofpro-inflammatory cytokines.

In some embodiments, the ECM composition comprises at least onepharmacological agent or composition (or drug), i.e. an agent orcomposition that is capable of producing a desired biological effect invivo, e.g., stimulation or suppression of apoptosis, stimulation orsuppression of an immune response, etc.

Suitable pharmacological agents and compositions include any of theaforementioned agents, including, without limitation, antibiotics,anti-fibrotics, anti-viral agents, analgesics, steroidalanti-inflammatories, non-steroidal anti-inflammatories,anti-neoplastics, anti-spasmodics, modulators of cell-extracellularmatrix interactions, proteins, hormones, enzymes and enzyme inhibitors,anticoagulants and/or anti-thrombotic agents, DNA, RNA, modified DNA andRNA, NSAIDs, inhibitors of DNA, RNA or protein synthesis, polypeptides,oligonucleotides, polynucleotides, nucleoproteins, compounds modulatingcell migration, compounds modulating proliferation and growth of tissue,and vasodilating agents.

In some embodiments of the invention, the pharmacological agentcomprises one of the aforementioned anti-inflammatory agents.

According to the invention, the wt. % of the pharmacological agent inthe ECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is dependent on thepharmacological agent(s) employed in the composition.

By way of example, as discussed in detail below, when thepharmacological agent comprises an antibiotic, the wt. % of theantibiotic in the ECM composition (and ECM-mimicking, ECM/ECM-mimicking,and statin augmented compositions of the invention) is in the range ofapproximately 0.0001-20 wt. %, more preferably, in the range ofapproximately 0.001-1 wt. %

In a preferred embodiment, the wt. % of the pharmacological agent in theECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is sufficient to induce ormodulate a desired biological effect in vivo, e.g., regeneration andremodeling of damaged biological tissue, without inducing an adversebiological response.

In some embodiments of the invention, the pharmacological agentcomprises a statin, i.e. a HMG-CoA reductase inhibitor. According to theinvention, suitable statins include, without limitation, atorvastatin(Lipitor®), cerivastatin, fluvastatin (Lescol®), lovastatin (Mevacor®,Altocor®, Altoprev®), mevastatin, pitavastatin (Livalo®, Pitava®),pravastatin (Pravachol®, Selektine®, Lipostat®), rosuvastatin(Crestor®), and simvastatin (Zocor®, Lipex®). Several actives comprisinga combination of a statin and another agent, such asezetimbe/simvastatin (Vytorin®), are also suitable.

In some embodiments, the wt. % of the HMG-CoA reductase inhibitor in theECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is in the range ofapproximately 0.0001-0.2 wt. %, more preferably, in the range ofapproximately 0.01-0.1 wt. %.

In a preferred embodiment, the wt. % of the HMG-CoA reductase inhibitorin the ECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is similarly sufficient toinduce or modulate a desired biological effect in vivo, e.g.,regeneration and remodeling of damaged biological tissue, withoutinducing an adverse biological response.

According to the invention, when the ECM composition and, hence,cardiovascular prostheses formed therefrom, is disposed proximatedamaged or diseased biological tissue, “modulated healing” iseffectuated.

The term “modulated healing”, as used herein, and variants of thislanguage generally refer to the modulation (e.g., alteration, delay,retardation, reduction, etc.) of a process involving different cascadesor sequences of naturally occurring tissue repair in response tolocalized tissue damage or injury, substantially reducing theirinflammatory effect. Modulated healing, as used herein, includes manydifferent biologic processes, including epithelial growth, fibrindeposition, platelet activation and attachment, inhibition,proliferation and/or differentiation, connective fibrous tissueproduction and function, angiogenesis, and several stages of acuteand/or chronic, i.e. wound healing, inflammation, and their interplaywith each other.

In such an instance, a minor amount of inflammation may ensue inresponse to tissue injury, but this level of inflammation response,e.g., platelet and/or fibrin deposition, is substantially reduced whencompared to inflammation that takes place in the absence of an ECMcomposition of the invention.

By way of example, in some embodiments, an ECM composition (and/orECM-mimicking composition and/or ECM/ECM-mimicking composition,discussed in detail below) and, hence, cardiovascular prosthesis formedtherefrom, of the invention is specifically formulated (or designed) toalter, delay, retard, reduce, and/or detain one or more of the phasesassociated with healing of damaged biological tissue, including, but notlimited to, the inflammatory phase (e.g., platelet or fibrindeposition), proliferative phase and maturation phase.

In some embodiments, “modulated healing” refers to the ability of an ECMcomposition (and/or ECM-mimicking composition and/or ECM/ECM-mimickingcomposition) and, hence, cardiovascular prosthesis formed therefrom, ofthe invention to alter a substantial inflammatory phase (e.g., plateletor fibrin deposition) at the beginning of the tissue healing process. Asused herein, the phrases “alter a substantial inflammatory phase”,“modulate inflammation” and “inflammation modulation” refer to theability of an ECM composition to substantially reduce an adverseinflammatory response at an injury site and induce “wound healing”,immune responses.

In some embodiments, the term “modulated healing” also refers to theability of an ECM composition (and/or ECM-mimicking composition and/orECM/ECM-mimicking composition) and, hence, cardiovascular prosthesisformed therefrom, of the invention to modulate inflammation of damagedbiological tissue by reducing the infiltration of “acute inflammatory”M1 macrophages and increasing the migration and, hence, population of“wound healing” M2 macrophages.

In some embodiments of the invention, “modulated healing” refers to theability of an ECM composition (and/or ECM-mimicking composition and/orECM/ECM-mimicking composition) and, hence, cardiovascular prosthesisformed therefrom, of the invention to induce neovascularization,including vasculogenesis, angiogenesis, and intussusception, host celland/or tissue proliferation, remodeling of damaged biological tissue,and regeneration of new tissue and tissue structures with site-specificstructural and functional properties.

As indicated above, in some embodiments of the invention, an ECMcomposition comprises a statin, i.e. a HMG-CoA reductase inhibitor.

The most preferred statin is cerivastatin, i.e.(3R,5S,6E)-7-[4-(4-fluorophenyl)-5-(methoxymethyl)-2,6-bis(propan-2-yl)py-ridin-3-yl]-3,5-dihydroxyhept-6-enoicacid.

According to the invention, when an ECM composition (and/or andECM/ECM-mimicking composition) comprising acellular ECM and a statin;particularly, cerivastatin (and/or an ECM-mimicking compositioncomprising a statin), i.e. a statin augmented composition, is disposed(i.e. delivered or administered) proximate damaged biological tissue,the statin augmented composition induces several beneficial biochemicalactions or activities, which enhance modulated healing.

The beneficial biochemical actions or activities induced when a statinaugmented composition is disposed to biological tissue; particularly,damaged cardiovascular tissue, are illustrated in FIG. 1.

Further details regarding the beneficial biochemical actions oractivities induced when a statin augmented composition is disposed tobiological tissue are set forth in U.S. Pat. No. 9,119,899, which isincorporated by reference herein in its entirety.

As discussed in detail below, one of the seminal biochemical actions oractivities induced by the statin augmented composition is highlyeffective inflammation modulation of damaged biological tissue when thestatin augmented composition is disposed proximate thereto.

As is well known in the art, damaged biological tissue undergoesinflammation-mediated repair in three phases: the inflammatory phase,proliferative phase and the maturation phase.

The inflammatory phase is an acute immune response where cytokinessignal the recruitment of leukocytes, including phagocytes, e.g. M1macrophages and dendritic cells, which clear dead endogenous or nativecells. Matrix metalloproteinase (MMP) expression also signals thebreakdown of endogenous ECM within the damaged biological tissue, whichoften occurs within ten (10) minutes after cardiovascular tissue damage.

The proliferative phase is a chronic immune response where transforminggrowth factor beta (TGF-β) and anti-inflammatory interleukin-10 (IL-10)suppress chemokine and inflammatory cytokine response, while promotingmyofibroblast cell proliferation. The myofibroblast cells then deposit aplurality of ECM proteins, which provide a “provisional” endogenous ECM.

The maturation phase is the scarring phase where endogenous ECM proteinsare crosslinked via lysyl oxidase and the myofibroblast cellssubsequently enter a quiescent state. The resulting accumulation ofcrosslinked “provisional” ECM (or fibrotic scar tissue) often results ina disruption of the endogenous ECM network.

The “provisional” ECM lacks the anisotropic network of endogenous ECMand, hence, is structurally weaker than endogenous ECM. By way ofexample, in cardiovascular tissue, the lack of an anisotropic networkdetrimentally alters the ventricular geometry, which leads to bothsystolic and diastolic dysfunction of the endogenous cardiovascular ECMand tissue structures.

One of the seminal MMPs activated during the inflammatory phase ismacrophage elastase or MMP-12, which directly breaks down the ECMprotein elastin and activates monocyte chemoattractant protein-1(MCP-1). MCP-1 recruits monocytes (macrophage and dendritic cellprogenitors), NK cells, T-lymphocytes, and dendritic cells to the sitesof inflammation. Further, the recruited M1 type macrophages secreteadditional MMP-12, thus creating a positive feedback loop thatperpetuates the inflammatory phase and breakdown of endogenous ECM.

As evidenced by the graphical illustrations shown in FIGS. 16-19, when astatin augmented composition is delivered to damaged biological tissue,the statin augmented composition inhibits expression of MCP-1, whichcloses the positive feedback loop that perpetuates the detrimentalbreakdown of endogenous ECM. The noted inhibition of MCP-1 expressionsubsequently abates the migration of pro-inflammatory cells, including,monocytes, M1 type macrophages, memory T-cells, and dendritic cells,which produces an anti-inflammatory effect.

When the statin augmented composition comprises acellular ECM andcerivastatin and the noted statin, i.e. cerivastatin, augmented ECMcomposition is disposed proximate damaged biological tissue, the statinaugmented ECM composition also inhibits expression of C—C chemokinereceptor type 2 (CCR2), which is the receptor protein for MCP-1. Thenoted restriction of both MCP-1 and CCR2 expression provides an enhancedlevel of inflammation modulation of damaged biological tissue when acerivastatin augmented ECM composition is disposed proximate thereto.

Thus, in some embodiments, the term “modulated healing” also refers tothe ability of an ECM composition (and ECM/ECM-mimicking composition);particularly, a cerivastatin augmented ECM composition to modulateinflammation by, among other actions, restricting expression of MCP-1and CCR2.

In some embodiments, “modulated healing” refers to the ability of an ECMcomposition (and/or ECM-mimicking composition and/or ECM/ECM-mimickingcomposition) and, hence, cardiovascular prosthesis formed therefrom, ofthe invention to induce anti-microbial and anti-biofilm activity, whichsignificantly enhance inflammation modulation of damaged biologicaltissue and, thereby, enhanced neovascularization, remodeling of thedamaged biological tissue and regeneration of new tissue and tissuestructures.

As also indicated above, in some embodiments, the ECM composition(and/or ECM-mimicking composition and/or ECM/ECM-mimicking composition)and, hence, cardiovascular prosthesis formed therefrom further comprisesan antibiotic. ECM, ECM-mimicking and ECM/ECM-mimicking compositionscomprising an antibiotic and hereinafter referred to as antibioticaugmented compositions.

According to the invention, when an antibiotic augmented compositionand, hence, cardiovascular prosthesis formed therefrom is delivereddirectly, i.e. local delivery, to damaged biological tissue, theantibiotic augmented composition induces several significant biologicalprocesses, including anti-microbial and anti-biofilm activity, which, asindicated above, significantly enhance modulated healing, includinginflammation modulation of the damaged biological tissue.

When an ECM composition (and/or an ECM/ECM-mimicking composition)comprising acellular ECM and, hence, cardiovascular prosthesis formedtherefrom is disposed proximate damaged biological tissue, migratingendogenous cells bind to the damaged biological tissue, whereby aplurality of acellular ECM components are degraded or “broken-down” bythe endogenous cells to form ECM bi-products. The ECM bi-productsprovide the “building-blocks” for regeneration of new tissue and tissuestructures.

A significant portion of the ECM bi-products comprise a particularsubtype of peptides known as anti-microbial peptides (AMPs), which are aseminal component of acellular ECM.

AMPs are a subtype of anti-microbial (and anti-biofilm) peptidemolecules, which provide a first line of defense against a plurality ofpathogens; particularly, microbes, e.g. bacteria.

AMPs comprise a net positive charge, which provides AMPs with theability to target and destroy microbes, such as bacteria, by disruptingthe cell membranes and/or walls of the microbes.

When the antibiotic augmented composition comprising acellular ECM, i.e.an antibiotic augmented ECM composition, is disposed proximate (i.e.delivered to) damaged biological tissue, the antibiotic augmented ECMcomposition releases the antibiotic and AMPs into the damaged biologicaltissue. As indicated above, the released AMPs disrupt the cell membranesand/or walls of microbes, while the antibiotic disrupts the microbes'capacity to synthesize and, hence, repair the cell membrane and/or wall.The protective cell membrane and/or wall of the microbes is thuspermanently destroyed and the microbes are rendered ineffective.

According to the invention, another seminal biochemical action oractivity induced by an antibiotic augmented ECM composition and, hence,cardiovascular prosthesis formed therewith is anti-biofilm activity.

Biofilm comprises a colony of bacteria that are embedded within aself-produced matrix of extracellular polymeric substance (EPS), whichcomprises a conglomeration consisting generally of extracellular DNA(eDNA), proteins, and polysaccharides.

The primary seminal function of a biofilm is to protect the bacterialcolonies from external assault; particularly, (i) anti-microbial agents,such as antibiotics and (ii) immune responses from a given host.

According to the invention, when an antibiotic augmented ECM compositionand, hence, cardiovascular prosthesis of the invention is disposedproximate damaged biological tissue that contains a bacterial biofilm,the antibiotic augmented ECM composition and, hence, cardiovascularprosthesis releases matrix metalloproteinases (MMPs), which provide twoseminal functions: (i) degrade a portion of the tissue during theremodeling and regeneration processes to form tissue bi-products and(ii) degrade the EPS that forms the bacterial biofilm.

As also indicated above, when an antibiotic augmented ECM composition isdisposed proximate damaged biological tissue, AMPs are released into thedamaged biological tissue. The cationic properties of the AMPs provideAMPs with the ability to bind to DNA, which has an inherently anionicnature.

The AMPs are thus able to bind to the eDNA component of bacterialbiofilms, which targets the eDNA for degradation by an endogenous immuneresponse in the damaged biological tissue. The AMPs are also capable ofdisrupting the attachment of bacteria to the bacterial biofilm, whichrenders the bacteria planktonic and, thus, significantly easier todestroy in vivo via antibiotics and/or an in vivo immune response.

In some embodiments of the invention, the antibiotic augmented ECMcomposition preferably comprises vancomycin and gentamicin.

Vancomycin and gentamicin have distinct binding affinities for acellularECM, which provides a “two-phase” antibiotic delivery profile: (i) a“bolus” antibiotic delivery profile and (ii) a sustained releaseantibiotic profile.

Vancomycin has a higher binding affinity for acellular ECM thangentamicin. Thus, according to the invention, when an antibioticaugmented ECM composition comprises vancomycin and gentamicin isdisposed proximate to damaged biological tissue, by virtue ofvancomycin's higher binding affinity for the acellular ECM, vancomycinis delivered to the damaged biological tissue at a substantially slowerrate than the gentamicin, i.e. a sustained release delivery profile.Since gentamicin's binding affinity for acellular ECM is substantiallylower than vancomycin's binding affinity, a bolus delivery of gentamicinto the damaged biological tissue is effectuated.

The initial bolus of gentamicin renders the microbes, e.g. bacteria, andbiofilm at the damaged biological tissue site ineffective, while thesustained delivery of vancomycin prevents the colonialization ofplanktonic microbes and the subsequent formation of biofilms.

In some embodiments of the invention, the wt. % of the antibiotic in theECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is in the range ofapproximately 0.0001-20 wt. %, more preferably, in the range ofapproximately 0.001-1 wt. %.

In some embodiments of the invention, the wt. % of the antibiotic in theECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is less than approximately0.0001 wt. %.

In a preferred embodiment, the wt. % of the antibiotic in the ECMcomposition (and ECM-mimicking, ECM/ECM-mimicking, and statin augmentedcompositions of the invention) is sufficient to induce severalsignificant biological processes, including anti-microbial andanti-biofilm activity in vivo, e.g., disrupting the cell membranesand/or walls of microbes, without inducing an adverse biologicalresponse.

As also indicated above, in some embodiments of the invention, the ECMcomposition (and/or ECM-mimicking composition and/or ECM/ECM-mimickingcomposition) and, hence, cardiovascular prosthesis formed therefromfurther comprises at least one additional biologically active agent orcomposition, i.e. an agent that induces or modulates a physiological orbiological process, or cellular activity, e.g., induces proliferation,and/or growth and/or regeneration of tissue.

ECM, ECM-mimicking and ECM/ECM-mimicking compositions comprising agrowth factor are hereinafter referred to as growth factor augmentedcompositions.

In some embodiments of the invention, the biologically active agentcomprises a growth factor selected from the group comprisingtransforming growth factor-alpha (TGF-α), transforming growthfactor-beta (TGF-β), basic fibroblast growth factor (bFGF), and vascularepithelial growth factor (VEGF).

According to the invention, when a growth factor augmented compositionand, hence, cardiovascular prosthesis formed therefrom is disposedproximate damaged tissue, the growth factors link to and interact withat least one molecule in the composition and further induce and/orcontrol modulated healing, i.e. inflammation modulation and/or host celland/or tissue proliferation, and/or remodeling of damaged biologicaltissue, and regeneration of new tissue and tissue structures.

According to the invention, when a growth factor augmented compositioncomprises VEGF and a second growth factor, comprising bFGF, TGF-α orTGF-β and the noted growth factor augmented composition is disposedproximate damaged biological tissue, synergistic activity by and betweenVEGF and the second growth factor is induced; the synergistic activitycomprising induced angiogenesis by VEGF, which facilitates cellproliferation and enhances bioremodeling of the damaged biologicaltissue induced by the second growth factor, i.e. bFGF, TGF-α or TGF-β.

In some embodiments of the invention, the biologically active agentcomprises an exosome. As indicated above, exosomes comprise a lipidbilayer structure that contains or encapsulates a biologically activeagent, such as a growth factor, e.g. TGF-β, TGF-α, VEGF and insulin-likegrowth factor (IGF-I), cytokine, e.g. interleukin-8 (IL-8),transcription factor and micro RNA (miRNA).

ECM, ECM-mimicking and ECM/ECM-mimicking compositions comprising anexosome are hereinafter referred to as exosome augmented compositions.

Exosomes significantly enhance the delivery of biologically activeagents to cells through two seminal properties/capabilities. The firstproperty comprises the capacity of exosomes to shield the encapsulatedbiologically active agents (via the exosome lipid bilayer) fromproteolytic agents, which can, and often will, degrade unshielded (orfree) bioactive molecules and render the molecules non-functional inbiological tissue environments.

The second property of exosomes comprises the capacity to directly and,hence, more efficiently deliver biologically active agents to endogenouscells in the biological tissue. As is well known in the art, endogenouscells typically do not comprise the capacity to “directly” interact with“free” biologically active agents, such as growth factors. There must beadditional biological processes initiated by the endogenous cells tointeract directly with biologically active agents, e.g. expression ofreceptor proteins for or corresponding to the biologically activeagents.

Exosomes facilitate direct interaction by and between endogenous cellsand exosome encapsulated biologically active agents (and, hence, directdelivery of bioactive molecules to endogenous cells), which enhances thebioactivity of the agents.

According to the invention, when an exosome composition comprisesacellular ECM and the exosome augmented composition is delivered to thedamaged biological tissue, the noted exosome augmented ECM composition“concomitantly” induces a multitude of significant biological processesin vivo, including (i) significantly enhanced inflammation modulation ofthe damaged biological tissue, (ii) induced neovascularization, (iii)induced stem cell proliferation, (iv) induced remodeling of the damagedbiological tissue, and (v) induced regeneration of new tissue and tissuestructures with site-specific structural and functional properties,compared to acellular ECM alone.

By way of example, when an exosome augmented ECM composition comprisingencapsulated IL-8 (and, hence, cardiovascular prosthesis formedtherefrom) is disposed proximate damaged biological tissue, the exosomeencapsulated IL-8 and, hence, tissue prosthesis modulates the transitionof M1 type “acute inflammatory” macrophages to M2 type “wound healing”macrophages initiated by the acellular ECM.

By way of further example, when an exosome augmented ECM compositioncomprising encapsulated miRNAs (and, hence, cardiovascular prosthesisformed therefrom) is disposed proximate damaged biological tissue, thecardiovascular prosthesis induce enhanced stem cell proliferation viathe delivery of exosome encapsulated miRNAs and transcription factors tothe damaged biological tissue, which signals the endogenous stem cellsto bind and/or attach to the acellular ECM and proliferate.

In some embodiments, the exosomes are derived and, hence, processed froman aforementioned tissue source. In some embodiments, the exosomes areprocessed and derived from a mammalian fluid composition including, butnot limited to blood, amniotic fluid, lymphatic fluid, interstitialfluid, pleural fluid, peritoneal fluid, pericardial fluid andcerebrospinal fluid.

In some embodiments, exosomes are derived and, hence, processed from invitro or in vivo cultured cells. According to the invention, exosomescan be derived from any one of the aforementioned cells, such asmesenchymal stem cells and hematopoietic stem cells.

In some embodiments, mesenchymal stem cells are cultured in a cellculture media under hypoxic conditions to induce a higher productionrate of exosomes.

In some embodiments, mesenchymal stem cells are cultured on anaforementioned acellular ECM, where the mesenchymal stem cells conditionthe acellular ECM by releasing exosomes, which bind to the ECMcomposition to form an exosome augmented ECM composition and/orECM/ECM-mimicking composition.

In some embodiments, the exosomes comprise semi-synthetically generatedexosomes. According to the invention, the semi-synthetically generatedexosomes can be derived from an exosome producing cell line.

By way of example, semi-synthetically generated exosomes can begenerated by incubating mesenchymal stem cells in a medium comprising apredetermined concentration of any one of the aforementionedbiologically active agents and/or pharmacological active agents and,after a predetermined period of time, removing the mesenchymal stemcells from the incubating medium and in vitro culturing usingconventional cell culture techniques. The cell culture media employedcan then be processed to isolate one or more exosome-encapsulatedbiologically active agents and/or pharmacological active agents.

According to the invention, the exosome-encapsulated biologically activeagents and/or pharmacological active agents can be isolated from thecell culture media using any known conventional method, such asultra-centrifugation.

According to the invention, the semi-synthetically generated exosomesmarkedly improve the efficacy of the aforementioned biologically activeagents and/or the pharmacological active agents by providing a means oftraversing the cell membrane of endogenous cells.

As indicated above, in some embodiments, the wt. % of the biologicallyactive agent in the ECM composition (and ECM-mimicking,ECM/ECM-mimicking, and statin augmented compositions of the invention,discussed in detail below) is in the range of approximately 0.0001-20wt. %, more preferably, in the range of approximately 0.001-1 wt. %.

In some embodiments, the wt. % of a biologically active and/orpharmacological agent in an ECM, ECM-mimicking and/or ECM/ECM-mimickingcomposition is preferably less than 0.001%, less than 0.01%, less than0.1%, less than 0.5%, less than 1%, less than 2%, less than 5%, lessthan 10.

In some embodiments, the wt. % of a biologically active and/orpharmacological agent in an ECM, ECM-mimicking and/or ECM/ECM-mimickingcomposition is preferably greater than 0.0001%, greater than 0.001%,greater than 0.01%, greater than 0.1%, greater than 0.5%, greater than1%, greater than 1.5%, greater than 2%, greater than 4%, greater than5%, greater than 10%, greater than 12%, greater than 15%, and greaterthan 20%.

In a preferred embodiment, the wt. % of the biologically active agent inthe ECM composition (and ECM-mimicking, ECM/ECM-mimicking, and statinaugmented compositions of the invention) is sufficient to induce ormodulate a physiological or biological process, without inducing anadverse biological response, e.g., a physiological response that issufficient to induce a biological process and/or restrict a phaseassociated with biological tissue healing in vivo.

According to the invention, any of the compositions and, hence,cardiovascular prostheses referenced herein, such as an ECM composition,are configured to provide a single-stage agent delivery profile, i.e.comprise a single-stage delivery vehicle, wherein a modulated dosage ofa biologically active and/or pharmacological agent is provided. In someembodiments, the compositions provide a multi-stage agent deliveryprofile, i.e. comprise a multi-stage agent delivery vehicle, wherein aplurality of biologically active and/or pharmacological agents areadministered via a modulated dosage. Suitable single-stage andmulti-stage agent delivery vehicles are disclosed in Co-Pending U.S.application Ser. Nos. 14/554,730, 14/957,995, 14/958,061 and 14/958,034,which are incorporated by reference herein.

In some embodiments, the cardiovascular prosthesis comprises anECM-mimicking composition comprising PGS.

In some embodiments, the ECM-mimicking composition further comprises atleast one of the aforementioned biologically active agents and/orpharmacological agents.

In some embodiments, the cardiovascular prostheses comprise anECM/ECM-mimicking composition comprising acellular ECM and PGS.

In some embodiments, the ECM/ECM-mimicking composition further comprisesat least one of the aforementioned biologically active agents and/orpharmacological agents.

As discussed in detail below, PGS provides numerous beneficialstructural and biochemical actions or activities.

PGS Physical Properties

PGS is a condensate of the non-immunogenic compositions glycerol (asimple sugar alcohol) and sebacic acid (a naturally occurringdicarboxylic acid), wherein, glycerol and sebacic acid are readilymetabolized when disposed proximate mammalian tissue. Thenon-immunogenic properties substantially limit the acute inflammatoryresponses typically associated with other “biocompatible” polymers, suchas ePTFE (polytetrafluoroethylene), that are detrimental tobioremodeling and tissue regeneration.

As set forth in Co-pending U.S. application Ser. No. 14/566,359, whichis incorporated by reference herein, the tensile strength of the PGS isat least 0.28±0.004 MPa. The Young's modulus and elongation of PGS areat least 0.122±0.0003 and at least 237.8±0.64%, respectively.

Thus, according to the invention, when a cardiovascular prosthesis ofthe invention comprises PGS, i.e. formed from a composition comprisingPGS or includes a PGS layer or coating, the PGS enhances the mechanicalstrength of the prosthesis.

PGS Adhesive Properties

PGS also exhibits unique adhesive properties.

Thus, according to the invention, when a cardiovascular prosthesiscomprises a PGS layer and/or PGS coated surface that is in contact withbiological tissue, such as tissue of a cardiovascular structure, theprosthesis adheres thereto, which facilitates modulated healing by theprosthesis.

When a cardiovascular prosthesis comprises a PGS layer and/or PGScoated. surface that is in contact with a non-coated or PGS coatedsurface of a second prosthesis structure, e.g. a second sheet member ofa laminate structure, the PGS layer and/or PGS coated surface alsoadheres to the non-coated or PGS coated surface of the second prosthesisstructure, e.g., second sheet member, which substantially reduces oreliminates dilation and/or delamination of the prosthesis structure ormembers.

ECM-Mimicking Properties/Actions

PGS also induces tissue remodeling and regeneration when administered todamaged tissue, thus, mimicking the seminal regenerative properties ofacellular ECM and, hence, a composition formed therefrom. The mechanismunderlying this behavior is deemed to be based on the mechanical andbiodegradation kinetics of the PGS. Sant, et al., Effect ofBiodegradation and de novo Matrix Synthesis on the Mechanical Propertiesof VIC-seeded PGS-PCL scaffolds, Acta. Biomater., vol. 9(4), pp. 5963-73(2013) and Wang, et al., In Vivo Degradation Characteristics ofPoly(Glycerol Sebacate), J. Biomed. Mat. Res. Part(A), Vol. (66)1, pp.192-197 (2003), incorporated by reference herein.

When an ECM and/or ECM/ECM-mimicking composition comprising acellularECM and PGS is disposed proximate damaged tissue, such as damagedcardiovascular tissue, the synergistic action by and between PGS andacellular ECM provides an enhanced level of remodeling of the damagedtissue and regeneration of new tissue and tissue structures.

Degradation Modulation

PGS also modulates the degradation characteristics ECM andECM/ECM-mimicking compositions and structures formed therefrom.

By way of example, when a cardiovascular prosthesis comprises a PUSlayer or PGS coated outer surface and the cardiovascular prosthesis isdisposed proximate damaged biological tissue, e.g. damagedcardiovascular tissue, the PGS layer or PGS coated outer surfacemodulates degradation of the prosthesis.

In some embodiments, the cardiovascular prosthesis exhibits a lineardegradation profile, which induces an enhanced level of modulatedhealing, including remodeling of damaged biological tissue andregeneration of new tissue and tissue structures.

In some embodiments, the linear degradation profile provided by the PGSin an ECM or ECM/ECM-mimicking composition allows the cardiovascularprosthesis formed therefrom to degrade at a rate that reduces theprobability of maladaptive remodeling, e.g. fibrosis. Thus, the uniquespatial properties of the combined acellular ECM and PGS components of acardiovascular prosthesis further enhance modulated healing of damagedtissue.

In some embodiments of the invention, the ECM-mimicking compositioncomprises PGS and poly(ε-caprolactone) (PCL). According to theinvention, the addition of PCL to the ECM-mimicking biomaterialcomposition enhances the structural integrity of the cardiovascularprostheses and further modulates the degradation of a cardiovascularprostheses formed therefrom.

In some embodiments, the ECM-mimicking composition comprisespoly(glycerol sebacate) acrylate (PGSA), which, according to theinvention, can be crosslinked, i.e. cured, via the combination of aphotoinitiator and/or radiation.

According to the invention, suitable photoinitiators for radiationinduced crosslinking comprise, without limitation,2-hydroxy-1-[4-hydroxyethoxy) phenyl]-2-methyl-1-propanone (D 2959, CibaGeigy), 2,2-dimethoxy-2-phenylacetophenone, titanocenes, fluorinateddiaryltitanocenes, iron arene complexes, manganese decacarbonyl, methylcyclopentadienyl manganese tricarbonyl and any organometallaticphotoinitiator that produces free radicals or cations.

According to the invention, suitable radiation wavelengths forcrosslinking and/or curing the ECM-mimicking composition comprise,without limitation, visible light; particularly, radiation in the rangeof approximately 380-750 nm, and ultraviolet (UV) light, particularly,radiation in the range of 10-400 nm, which includes extreme UV (10-121nm), vacuum UV (10-200 nm), hydrogen lyman α-UV (121-122 nm), Far UV(122-200 nm), Middle UV (200-300 nm), Near UV (300-400 nm), UV-C(100-280 nm), UV-B (280-315 nm) and UV-A (315-400 nm) species of UVlight.

In some embodiments, the ECM-mimicking composition comprises aco-polymer of PGSA and polyethylene glycol (PEG) diacrylate.

In some embodiments, when a cardiovascular prosthesis is disposedproximate damaged biological tissue, modulated healing is effectuatedthrough the structural features of the cardiovascular prosthesis. Thestructural features of the cardiovascular prosthesis provide the spatialand mechanical cues to modulate endogenous cell polarity and alignment.The structural features of the cardiovascular prosthesis furthermodulate endogenous cell proliferation, migration and differentiation.

As discussed in detail above, the cardiovascular prostheses of theinvention can comprise various structures and compositions, including,but not limited to, particulate structures, mesh constructs, encasementstructures, coated structures and multi-sheet laminate structures.

Exemplar cardiovascular prostheses of the invention will now bedescribed in detail. It is, however, understood that the invention isnot limited to the structures described below. Indeed, as indicatedabove, the cardiovascular prostheses of the invention can comprisevarious structures and compositions.

As also indicated above, the cardiovascular prostheses can be employedto treat various disorders, including, without limitation, atrialfibrillation (pre- and post-operative) and the root causes thereof,damaged or diseased biological tissue, e.g., infarct tissue, damaged anddiseased mammalian organs and structures, e.g., infarct tissue, anddamaged and diseased mammalian organs and structures, e.g., cardiacvessels and valves, myocardium, pericardium, arteries, trachea, etc.esophagus

Sheet Structures

In some embodiments of the invention, the cardiovascular prosthesescomprise or are formed with sheet members.

Referring now to FIGS. 2 and 3, there is shown one embodiment of a sheetmember of the invention. As illustrated in FIGS. 2 and 3, the sheetmember 10 a comprises a top surface 14 and a bottom surface 12. In someembodiments of the invention, the top surface 14 defines a tissuecontacting surface.

In some embodiments, the sheet member 10 a comprises one of theaforementioned ECM compositions.

In some embodiments, the sheet member 10 a comprises one of theaforementioned ECM-mimicking compositions.

In some embodiments, the sheet member 10 a comprises one of theaforementioned ECM/ECM-mimicking compositions.

As set forth in Co-Pending application Ser. No. 14/566,306, which isincorporated by referenced herein, in some embodiments, at least onesurface 14, 12 of the sheet member 10 a comprises a crosslinked surface.In the illustrated embodiment, the top surface 14 comprises acrosslinked surface 16.

In some embodiments of the invention, the crosslinked surface 16comprises a chemically induced crosslinked surface.

In some embodiments of the invention, the crosslinked surface 16comprises an energy induced crosslinked surface.

According to the invention, the crosslinked surface 16 of the sheetmember 10 a is configured to adhere to biological tissue and/or a secondsheet member of a prosthesis structure, such as the laminate structuredescribed below, whereby dilation and/or delamination of the structureis substantially reduced or eliminated.

Referring now to FIGS. 4 and 5, there is shown another embodiment of asheet member of the invention. As illustrated in FIGS. 3 and 4, thesheet member 10 b similarly comprises bottom and top surfaces 12, 14.

In the illustrated embodiment, at least one surface 12, 14 of the sheetmember 10 b comprises an outer coating. In some embodiments, asillustrated in FIG. 4, the top surface 14 of the sheet member 10 bcomprises a coated surface or layer 18. In some embodiments, the coatedor layered top surface 14 similarly defines a tissue contacting surface.

In some embodiments, the coated surface or layer 18 comprises at leastone of the aforementioned ECM compositions.

In some embodiments, the coated surface or layer 18 comprises at leastone of the aforementioned ECM-mimicking compositions.

In some embodiments, coated surface or layer 18 comprises at least oneof the aforementioned ECM/ECM-mimicking compositions.

In some embodiments of the invention, the sheet members 10 a, 10 band/or coated surface or layer 18 further comprise at least one of theaforementioned biologically active agents or compositions.

In some embodiments of the invention, the ECM sheet members 10 a, 10 band/or coated surface or layer 18 further comprise at least one of theaforementioned pharmacological agents or compositions.

According to the invention, the ECM sheet members 10 a, 10 b can beemployed to construct various cardiovascular prosthesis structures,including, without limitation, single sheet structures, e.g. grafts,such as described in U.S. Pat. No. 8,877,224, and multi-sheetstructures, such as described in Co-Pending application Ser. Nos.14/566,359, 14/953,548 and 14/566,306. The noted applications areincorporated by reference herein in their entirety.

The single and multi-sheet structures can also comprise various shapesand dimensions to accommodate various applications.

Referring now to FIGS. 6-9, there are shown two (2) embodiments ofmulti-sheet prosthesis structures of the invention. Referring to FIGS. 6and 8, there is shown the multi-sheet prosthesis structures in apre-lamination configuration (denoted 20 a, 20 c). As illustrated inFIGS. 6 and 8, the multi-sheet structures comprise three (3) sheetmembers 10 c, 10 d, 10 e and 10 f, 10 g, 10 h.

According to the invention, the multi-sheet prosthesis structures canalso comprise less or more than three (3) sheet members, e.g., two (2)sheet members, five (5) sheet members, etc.

As illustrated in FIG. 6, in some embodiments of the invention, thefirst and second sheet members 10 c, 10 d comprise a top crosslinkedsurface 16 that is configured to adhere to the bottom surface 12 of theadjoining sheet members 10 d, 10 e to form the laminate structure shownin FIG. 7. structure with a non-crosslinked top and bottom surface.

As illustrated in FIG. 8, in some embodiments of the invention, thefirst and second sheet members 10 f, 10 g comprise a biomaterial coatedsurface 18 that is similarly configured to adhere to the bottom surface12 of the adjoining sheet members 10 g, 10 h to form the laminatestructure shown in FIG. 9.

In some embodiments of the invention, the biomaterial coated surface 18comprises one of the aforementioned ECM-mimicking compositions.

In some embodiments of the invention, the biomaterial coated surface 18comprises one of the aforementioned ECM/ECM-mimicking compositions.

As discussed in detail above, the biomaterial coated surface 18 is alsoconfigured to (i) adhere the multi-sheet structure 20 d to biologicaltissue and (ii) modulate degradation of the multi-sheet structure 20 dwhen the multi-sheet structure 20 d is in contact with biologicaltissue.

According to the invention, the ECM sheet members 10 a, 10 b can beemployed to form an encasement structure having a cavity therein that isconfigured to receive and, hence, encase a medical device and/or any oneof the aforementioned ECM, ECM-mimicking or ECM/ECM-mimickingcompositions and/or biologically active or pharmacological agents.

According to the invention, the encasement structures can comprisevarious shapes and sizes to accommodate virtually all shapes and sizesof medical devices and quantities of compositions.

Illustrative are the encasement structures described in U.S. Pat. Nos.8,758,448, 9,066,993, 9,333,277 and 9,283,302 and Co-Pending U.S.application Ser. Nos. 14/818,757, 14/819,964, 14/571,639, 14/571,679,14/685,755, 14/833,327, 14/833,340, 14/833,354, 14/833,373 and14/833,404, which are incorporated by reference herein in theirentirety.

Referring now to FIGS. 10-12, two (2) embodiments of encasementstructures will be described in detail.

Referring first to FIG. 10, there is shown an embodiment of anencasement structure 30 a in a folded, pre-lamination configuration. Asillustrated in FIG. 9, the encasement structure 30 a preferablycomprises one (1) sheet member 10 i.

In some embodiments of the invention, sheet member 10 i comprises sheet10 a shown in FIGS. 1 and 2 (denoted 10 i). According to the invention,the sheet member 10 i can also comprise sheet member 10 b shown in FIGS.4 and 5.

According to the invention, more than one (1) sheet member 10 i can beemployed to construct the encasement structure 30 a (and 30 b discussedbelow), wherein a multi-sheet encasement structure is provided.

As illustrated in FIGS. 10 and 11, the encasement structure 30 acomprises a top surface 14, sides 34 a, 34 b, and edge regions 32 a, 32b.

In some embodiments of the invention, at least one (1), preferably, bothsides 34 a, 34 b are laminated to form a pouch structure having a cavity40 therein that is preferably configured to encase a medical device 100therein.

As indicated above, in some embodiments of the invention, sheet member10 i comprises sheet member 10 b shown in FIGS. 4 and 5, comprising anECM-mimicking or ECM/ECM-mimicking coated surface 18. In suchembodiments, when the sheet member 10 i is folded over the coatedsurface (wherein the coated surface 18 forms or defines the encasementstructure cavity 40), the sides 34 a, 34 b adhere and seal theencasement structure about sides 34 a, 34 b.

Referring now to FIG. 12, there is shown another embodiment of anencasement structure 30 b. As illustrated in FIG. 12, the encasementstructure 30 b preferably comprises two (2) ECM sheet members 10 j, 10 kthat are joined on at least one end 36 a, 36 b. According to theinvention, the end or ends 36 a, 36 b can similarly be joined bylaminating the end or ends 36 a, 36 b or, as described above, employingat least one sheet member comprising an ECM-mimicking orECM/ECM-mimicking coated surface

Mesh Structures

According to the invention, the cardiovascular prostheses can alsocomprise mesh constructs comprising at least one biodegradable fiber. Insome embodiments, the cardiovascular prostheses comprise a plurality ofbiodegradable fibers, such as described in Co-Pending U.S. applicationSer. Nos. 14/554,730, 14/957,995 and 14/958,034, which are incorporatedby reference herein.

According to the invention, the biodegradable fibers can be arranged ororiented in various configurations, i.e. mesh patterns, to form a meshfiber member or construct, such as shown in FIG. 13.

Referring now to FIG. 13, in some embodiments of the invention, the meshconstructs 50 comprise a plurality of substantially perpendicularinterwoven or intersecting biodegradable fibers 52 contained by arestraining edge 54.

In some embodiments, the biodegradable fiber comprises at least one ofthe aforementioned ECM compositions.

In some embodiments, the biodegradable fiber comprises at least one ofthe aforementioned ECM-mimicking compositions.

In some embodiments, the biodegradable fiber comprises at least one ofthe aforementioned ECM/ECM-mimicking compositions.

In some embodiments, the biodegradable fiber comprises at least one ofthe aforementioned biologically active and/or pharmacological agents.

According to the invention, the mesh constructs can comprise anycombination of ECM, ECM-mimicking and/or ECM/ECM-mimicking compositionfibers.

According to the invention, the mesh constructs can also comprisebiodegradable fibers comprising different compositions and/ormulti-composition fibers, e.g., coated fibers.

Particulate Structures

According to the invention, the cardiovascular prostheses can alsocomprise mesh particulate structures, such as described in U.S. Pat.Nos. 9,072,816, 9,119,899, 8,962,324 and 8,568,761 and Co-Pending U.S.application Ser. No. 14/566,404, which are incorporated by referenceherein in their entirety.

According to the invention, the particulate structures can comprise anyof the aforementioned ECM, ECM-mimicking and/or ECM/ECM-mimickingcompositions and/or a mixture thereof.

Referring now to FIG. 14, in some embodiments of the invention, theparticulate structures comprise a core 62 and outer layer (or coating)64, such as described Co-Pending U.S. application Ser. Nos. 14/832,109and 14/832,163, which are incorporated by reference herein in theirentirety.

According to the invention, the core and/or outer layer 64 can similarlycomprise any of the aforementioned ECM, ECM-mimicking and/orECM/ECM-mimicking compositions and/or a mixture thereof.

In some embodiments of the invention, the outer layer comprises anECM-mimicking and/or ECM/ECM-mimicking composition. According to theinvention, when the particulate structure outer layer comprises anECM-mimicking and/or ECM/ECM-mimicking composition, the outer layer (i)enhances the structural integrity of the particulate structure and (ii)modulates the degradation characteristics of the particulate structurewhen disposed proximate biological tissue.

As described in Co-Pending U.S. application Ser. No. 14/832,109, variousconventional means can be employed to form a particulate structure ofthe invention.

In some embodiments of the invention, the cardiovascular prosthesescomprise a plurality of the particulate structure. According to theinvention, the particulate structures can be in the form of mixedliquids, mixed emulsions, mixed gels, mixed pastes, or mixed solidparticulates. The liquid or semi-solid components of the particulatecompositions can also comprise various concentrations.

EXAMPLES

The following examples are provided to enable those skilled in the artto more clearly understand and practice the present invention. Theyshould not be considered as limiting the scope of the invention, butmerely as being illustrated as representative thereof.

Example 1 Assessment of the Physiological Effects of a CerivastatinAugmented Sheet Member in a Canine Model

A study was performed using a canine model in order to evaluate thephysiological effects of various concentrations of cerivastatin in asheet member comprising acellular ECM derived from small intestinesubmucosa (SIS), i.e. CorMatrix® acellular ECM patches, at 2 and 24 hrspost-implantation in canine myocardium.

The study included a total of four (4) treatment groups consisting oftwo (2) canines per treatment group. The four treatment groups consistedof a control group where two (2) canines were treated with an ECM sheetmember derived from SIS without cerivastatin. The remaining three (3)treatment groups comprised groups of two (2) canines treated withcerivastatin augmented sheet members having 0.1 mg, 0.3 mg and 1 mg ofcerivastatin per ECM sheet member.

The cerivastatin was impregnated into a 9 cm×9 cm sheet member byincubating the sheet member in a solution of phosphate buffered saline(PBS) and cerivastatin for 24 hours.

After the sheet members were prepared and impregnated with cerivastatin,eight (8) canines were prepared for surgery. The canines receivedanesthetic premedication, including 1 mg of Atropine and 0.15 mg ofburenorphine, followed by anesthetic induction with propofol 100-300 mgor Pentothal 250-500 mg via intravenous injection. After the canineswere anesthetized, a baseline blood sample and a pericardium fluidsample were collected from all eight (8) of the canine subjects.

A sternotomy was performed on all eight (8) of the canine subjects. Thepericardial sac was isolated and a section of the pericardium wasexcised. A cerivastatin augmented sheet member was sutured to theexcised region of the pericardium of six (6) of the canines as a closuredevice. The sheet members without cerivastatin were sutured to theexcised region of the pericardium as a closure device for the remainingtwo (2) canines.

After 2 hours, a blood serum sample was collected from four (4) of thecanines. After 24 hours, a blood serum sample was collected from theremaining four (4) canines. Cardiac tissue samples were subsequentlyharvested from the hearts of the canines, cut in half, placed incryotubes, and flash frozen in liquid nitrogen. Samples were stored in a−80° C. freezer until used.

The tissue gene expression of MCP-1 and CCR2 were determined usingreverse transcription polymerase chain reaction (RT-PCR).

FIGS. 15 and 16 show CCR2 and MCP-1 mRNA/18s concentrations for thecardiac tissue samples harvested at the 2 hr. time point, compared tothe CCR2 and MCP-1 mRNA/18s concentrations for the tissue samplesharvested at the 24 hour time point, respectively.

As reflected in FIGS. 15 and 16, when a cerivastatin augmented sheetmember of the invention (and, hence, prosthesis structure formedtherewith), is disposed proximate to damaged cardiovascular tissue, thecerivastatin augmented sheet member restricts expression of MCP-1 andCCR2 and, thereby, modulates inflammation of the damaged cardiovasculartissue.

Example 2 Anti-Inflammatory Activity of Cerivastatin Delivered with anCerivastatin Augmented Sheet Member

In the following study, the activity of cerivastatin release from acerivastatin augmented sheet member comprising acellular ECM derivedfrom SIS was assessed using an in vitro transwell assay. Human monocyticcells, i.e. THP-1 cells, were seeded by preparing a solution of THP-1cells at 6×10⁵ cells/ml and loading the solution of THP-1 cells into thebottom of 12-well transwell plates.

The transwell assay included four (4) treatment groups, including acontrol group consisting of untreated THP-1 cells, a first control groupconsisting of THP-1 cells treated with 200 ng/ml lipopolysaccharide(LPS) alone, a second control group consisting of THP-1 cells treatedwith 200 ng/ml LPS and an ECM sheet member comprising SIS derived fromSIS, and a treatment group consisting of THP-1 cells treated with 200ng/ml LPS and a cerivastatin augmented ECM sheet member.

The THP-1 cells were treated with 200 ng/ml LPS to stimulate theproduction of MCP-1.

Over the course of four (4) days, seeded THP-1 cells were harvested oneach of the four (4) days of the study. Each of the harvested cellsamples were collected in a sealable vial and centrifuged to separatethe liquid media from the THP-1 cells. The liquid media, i.e.supernatant, was collected for use in an enzyme-linked immunosorbentassay. The THP-1 cells were also collected and used for RNA extractionand quantitative PCR.

FIGS. 17 and 18 reflect that treatment with 200 ng/ml of LPS inducedexpression of MCP-1 in THP-1 cells, and the placement of the sheetmember without cerivastatin on the seeded THP-1 cells did not produce asignificant change in MCP-1 expression. Conversely, the placement of thecerivastatin augmented sheet member on the seeded THP-1 cells restrictedMCP-1 expression. The restriction of MCP-1 was maintained over theentire course of the four (4) day experiment.

The results from the transwell assay further confirm that a cerivastatinaugmented sheet member (and, hence, cardiovascular prosthesis formedtherewith) is capable of modulating inflammation by restricting MCP-1expression.

Example 3 Determination of In Vitro Anti-Inflammatory Activity ofCerivastatin Released from a Cerivastatin Augmented Sheet Member

In the following study the activity of cerivastatin release from acerivastatin augmented sheet member comprising acellular ECM derivedfrom SIS was similarly assessed using the human monocytic cell lineTHP-1.

The THP-1 cells were seeded by preparing a solution of THP-1 cells at6×10³ cells/ml and loading the solution of THP-1 cells onto a cellculture plate.

The in vitro assay included six (6) treatment groups, including a firstcontrol group consisting of untreated THP-1 cells, a second controlgroup consisting of THP-1 cells treated with 200 ng/mllipopolysaccharide (LPS) alone, treatment groups consisting of THP-1cells treated with 200 ng/ml LPS and 0.1 μM of cerivastatin, 200 ng/mlLPS and 0.5 μM of cerivastatin, 200 ng/ml LPS and 1 μM of cerivastatinand 200 ng/ml LPS and 5 μM of cerivastatin.

The THP-1 cells were similarly treated with 200 ng/ml LPS to stimulatethe production of MCP-1.

Each of the cell samples were harvested and collected in a sealable vialand centrifuged to separate the liquid media from the THP-1 cells. TheTHP-1 cells were collected and used for RNA extraction and quantitativePCR.

As reflected in FIG. 19, the expression of MCP-1 mRNA was inhibited bycerivastatin in a concentration-dependent manner, i.e. higherconcentrations of cerivastatin resulted in greater restriction of MCP-1expression.

Example 4 Determination of MCP-1 and CCR2 Restriction by CerivastatinCompared to Other Statins

In the following study, the activity of cerivastatin and other statins,including simvastatin, lovastatin and atorvastatin, was similarlyassessed using the human monocytic cell line THP-1.

The THP-1 cells were similarly seeded by preparing a solution of THP-1cells at 6×10⁵ cells/ml and loading the solution of THP-1 cells onto acell culture plate.

The in vitro assay included six (6) treatment groups, including one (1)control group consisting of untreated THP-1 cells, one (1) positivecontrol group consisting of THP-1 cells treated with tumor necrosisfactor alpha (TNF-α) alone, four (4) treatment groups consisting ofTHP-1 cells treated with TNF-α and cerivastatin, TNF-α and simvastatin,TNF-α and lovastatin and TNF-α and atorvastatin.

In this study, the THP-1 cells were treated with TNF-α to stimulate theproduction of CCR2.

Each of the six (6) THP-1 cell samples were harvested and collected in asealable vial and centrifuged to separate the liquid media from theTHP-1 cells. The THP-1 cells were then collected and used for RNAextraction and quantitative PCR.

It was found that cerivastatin was capable of substantially restrictingexpression of CCR2. Simvastatin, lovastatin and atorvastatin did not,however, restrict CCR2 expression at levels comparable to cerivastatin.

Without departing from the spirit and scope of this invention, one ofordinary skill can make various changes and modifications to theinvention to adapt it to various usages and conditions. As such, thesechanges and modifications are properly, equitably, and intended to be,within the full range of equivalence of the following claims.

What is claimed is:
 1. A vascular graft for treating damaged or diseasedtissue, comprising: a sheet member comprising top and bottom surfaces,and an outer coating layer disposed on said sheet member top surface,said outer coating layer comprising poly(glycerol sebacate) (PGS), saidsheet member consisting of an exosome augmented composition consistingof acellular ECM derived from mammalian tissue source selected from thegroup consisting of small intestine submucosa (SIS), mesothelial tissue,placental tissue and cardiac tissue and a plurality of exogenousexosomes derived from mesenchymal stem cells (MSCs), said sheet member,when disposed proximate damaged tissue, being adapted to reduce aninflammatory phase of said damaged tissue and, thereby, reduce aninflammatory response thereof, whereby said sheet member inducesenhanced, stem cell proliferation, neovascularization, remodeling ofsaid damaged tissue, and regeneration of new tissue and tissuestructures with site specific structural and functional properties,compared to induced stem cell proliferation, neovascularization,remodeling, and regeneration of new tissue and tissue structures by anECM sheet member consisting solely of acellular mammalian ECM, saidcoated top surface of said sheet member, when disposed proximate saiddamaged tissue, being adapted to adhere to said damaged tissue, degradeand exhibit a linear degradation profile.